Mechanosensitive ion channels play a critical role in transducing physical stresses at the cell membrane into an electrochemical response. The MscL family of mechanosensitive channels is widely distributed among prokaryotes, and may participate in the regulation of osmotic pressure changes within the cell. In an effort to better understand the structural basis for the function of these channels, we have determined the structure of the MscL homologue from Mycobacterium tuberculosis by x-ray crystallography to 3.5 E resolution using diffraction data collected primarily at SSRL. This channel is organized as a homopentamer with each subunit containing two transmembrane ?-helices and a third cytoplasmic ?-helix. From the extracellular side, an ~18 E diameter water-filled opening leads into a funnel-shaped pore lined with hydrophilic residues which narrows at the cytoplasmic side to an occluded hydrophobic apex that may act as the channel gate. This structure may serve as a model for other mechanosensitive channels, as well as the broader class of pentameric ligand-gated ion channels exemplified by the nicotinic acetylcholine receptor.